Ken-ichiro Mutoh

Isolation, Identification, and Plaque Titration of Parvovirus from Muscovy Ducks in Japan

Muscovy ducks (Cairina moschata) showed abnormal feathering, leg weakness, and high mortality. A virus was isolated from these ducks after several blind passages in embryonating Muscovy duck eggs. The isolate was resistant to chloroform, to pH 3.2, and to 65 C for 30 min. Electron microscopy showed that the isolate was an icosahedral and nonenveloped virus 20-22 nm in diameter. The isolate reacted with an antiserum against a goose parvovirus in agar gel precipitation tests. After 15 passages of the isolate in embryonating eggs, the isolate was adapted to Muscovy duck embryo fibroblasts. The adapted virus developed cytopathic effects and made clear plaques on sheets of the fibroblasts. When 5-iodo-2-deoxyuridine was added to the culture medium, virus growth was inhibited. From the data shown above, the isolate was identified as a goose parvovirus.

Centroacinar and intercalated duct cells as potential precursors of pancreatic endocrine cells in rats treated with streptozotocin.

The present study examined the possibility for regeneration of pancreatic endocrine cells from centroacinar (CA) and intercalated duct (ICD) cells in rat pancreas after 5 days of continuous streptozotocin (STZ) administration. Nine rats were divided into 3 experimental groups: 1) Control group, 2) Short term recovery group; three days after STZ administration (STZ 3), and 3) Long term recovery group; ten days post-STZ administration (STZ 10). The CA and ICD cells in the STZ 3 group had swollen cytoplasm, and sometimes contained a vesicle within the core. An insulin positive signal was detected in and around the CA and ICD cells. In the STZ 3 group, cytokeratin 20 signals were co-localized with insulin signals in both CA and ICD cells. Electron microscopically, endocrine cells and small pancreatic islets were in close contact with CA and ICD cells. Systemic biophysical serum data reflected these immunohistological results. The present results suggest that CA and ICD cells are involved in the regeneration of pancreatic B cells in rats following a lesion produced by five consecutive days of STZ administration.

Immunocytochemical localization of annexin 5, a calcium-dependent phospholipid-binding protein, in rat endocrine organs.

Abstract Annexin 5, a unique calcium- and phospholipid-binding protein, has been investigated for its specific distribution in rat endocrine organs by immunocytochemistry with a specific antiserum to recombinant rat annexin 5. Follicular epithelial cells and parafollicular cells of the thyroid gland, adrenocortical cells of the zona fasciculata and zona reticularis, luteal cells, testicular interstitial cells, and Sertoli cells were shown to contain annexin 5. To examine whether the synthesis of annexin 5 would be affected by a change in humoral signal, the distribution of annexin 5 in the anterior pituitary was examined three weeks after ovariectomy. The withdrawal of ovarian hormones induced huge castration cells in the anterior pituitary gland, which contained abundant annexin 5. Annexin 5 was not detected in the pineal gland, the parathyroid gland, the islet of Langerhans, the adrenal medulla, zona glomerulosa cells, and granulosa cells. Since annexin 5 was shown to exist in many of the endocrine tissues examined, to be localized in specific cell types, and to be abundant in castration cells, it is suggested that annexin 5 contributes to secretory cell functions, which may be common to endocrine cells secreting chemically different hormones.

Distributional changes of BrdU, PCNA, E2F1 and PAL31 molecules in developing murine palatal rugae

The distribution of cells incorporating bromodeoxyuridine (BrdU) and the expression of molecules involved in the control of cell proliferation (proliferating cell nuclear antigen [PCNA], a cellular factor in F9 teratocarcinoma cells that recognizes an adenovirus E1A inducible promoter 1 [E2F1] and proliferation-related acidic nuclear protein 31 [PAL31]) during morphogenesis of the murine palatine rugae (PR) was examined histochemically. Pattern formation of the PR rudiment was initiated with cell cycle related molecules in the epithelium of the primary palate. Cells which had incorporated BrdU were detected at the outer areas of the presumptive epithelial placode (EP) and the EP at 11.5-13.5 days post coitum (dpc) and the outer areas of the PR protrusion after 14.5 dpc. The number of PCNA-positive cells at the central area of the PR protrusion decreased after 16.5 dpc. E2F-positive cells were detected at the outer areas of the PR protrusion at 15.5 and 16.5 dpc. The number of PAL31-positive cells at the presumptive EP area and the already-formed EP area was decreased at 11.5-13.5 dpc. In two dimensional histological reconstructions, PAL31 expression approximately corresponded to the distribution of BrdU-positive cells at 11.5 and 13.5 dpc. EP placode formation might be regulated by spatiotemporal cell proliferation control involving the expression of the PAL31 molecule. Following EP formation, PR development and growth control involved the expression of E2F1 and PCNA molecules.

Activin A and Equine Chorionic Gonadotropin Recover Reproductive Dysfunction Induced by Neonatal Exposure to an Estrogenic Endocrine Disruptor in Adult Male Mice

Abstract We aimed to elucidate the mechanism of action of estrogenic endocrine disruptors and the rescue of reproductive function, particularly the responsiveness of testes to eCG and/or activin A (ACT) after establishing reproductive disorders. Newborn male mice (n = 29) were randomly divided into an untreated group and three treatment groups that received diethylstilbestrol (DES; 100 μg per animal) subcutaneously on Postnatal Day 3 to establish reproductive disorders and daily treatment with PBS (controls: DES + PBS), eCG (eCG group: DES + eCG), or eCG + ACT (eCG + ACT group: DES + eCG + ACT) at 6–8 wk of age prior to mating. After treatment, the controls showed diminished Leydig cells in the testes and thin germ cell layers containing pyknotic germ cells and multinucleated cells. In the eCG and eCG + ACT groups, spermatids and Leydig cells increased markedly. The immunoexpression of androgen receptors in the eCG group and steroidogenic acute regulatory (STAR) protein in the eCG and eCG + ACT groups recovered to approximately the levels in the untreated group; plasma LH and testosterone levels also increased relative to those in the controls. In addition, the cell proliferation index, which is estimated from 5-bromo-2′-deoxyuridine immunoexpression in spermatogonia, increased significantly under eCG treatment, and even more with eCG + ACT. However, the numbers of germ and Leydig cells decreased at 12 wk of age. Thus, ACT and eCG help the testes to recover from the dysfunction induced by neonatal DES administration. Furthermore, the permanent male reproductive disorder induced by neonatal exposure to estrogenic agents may be more likely to result from dysfunction of the hypothalamic-pituitary axis than from dysfunction of the lower reproductive organs.

A novel mechanism of autolysis in Helicobacter pylori : Possible involvement of peptidergic substances

Background.  Helicobacter pylori survival in a hostile acidic environment is known to be caused by its production of urease, which is not released by known secretion pathways. It has been proposed that H. pylori cells undergo spontaneous autolysis during cultivation and that urease becomes surface‐associated only concomitant with bacterial autolysis. The aim of this study was to elucidate mechanisms by which H. pylori cells undergo autolysis during cultivation.

Morphological Changes in the Rat Endocrine Pancreas within 12 h of Intravenous Streptozotocin Administration

We examined early morphological changes in pancreatic endocrine cells within 12 h of intravenous streptozotocin (STZ) administration (60 mg/kg). Thirty rats were allocated either to a control group (vehicle alone) or to one of four experimental groups tested after 3, 6, 9 and 12 h. Karyopyknosis and cytoplasmic vacuoles were first observed in β‐cell cytoplasm 3 h after STZ administration (STZ‐3 h), and the most severe damage was found in β cells at STZ‐12 h. Insulin‐positive non‐islet cells were observed near the intercalated duct (ICD) and/or centroacinar (CA) cells at STZ‐6 h and their numbers peaked at STZ‐6 h. The distribution patterns of the insulin‐positive cells and those of nestin and insulin‐like growth factor‐1 were similar and their nuclei were positive for proliferating cell nuclear antigen. Thus, ICD cells and/or CA cells reacted immediately to transform into insulin‐secreting cells to replace injured β cells (or to compensate for the lack of β cells) within 12 h of STZ administration.

Expression of Nestin and IGF-1 in Rat Pancreas after Streptozotocin Administration

The present study examines whether centroacinar (CA) and intercalated duct (ICD) cells can serve as stem cells, after administration of the diabetogenic agent streptozotocin (STZ). Thirty rats were divided into five experimental groups: (1) control, (2) 1 day after STZ (STZ‐1), (3) 3 days after STZ (STZ‐3), (4) 7 days after STZ (STZ‐7) and (5) 14 days after STZ (STZ‐14). Many small pancreatic islets were observed in the STZ‐7 group than in the other experimental groups, and many of these small islets were in close contact with ICD and CA cells. A higher number of nestin, insulin‐like growth factor‐1 (IGF‐1) and IGF‐1‐receptor positive ICD and CA cells were observed at STZ‐3 and STZ‐7 than at the others. These expression patterns coincided well with the proliferating cell nuclear antigen pattern. The results suggest that rat pancreatic endocrine cells after damage by STZ administration might be recovered from newly generated cells derived from ICD and CA cells.

Distribution of Aromatase and Sex Steroid Receptors in the Baculum During the Rat Life Cycle: Effects of Estrogen During the Early Development of the Baculum

The baculum, also called os penis, plays an important role during copulation. However, the hormonal regulation of its development remains to be elucidated. To determine the direct involvement of sex steroids in the development of the baculum of rats, the distributions of androgen receptors (ARs), aromatase, and estrogen receptor alpha (ESR1) were observed immunohistochemically. On Postnatal Day 1, the rudiment of the baculum expressed ARs, aromatase, and ESR1. In the proximal segment of the baculum of neonatal rats, ARs were expressed in the parosteal layer but not in the periosteum or osteoblasts. Aromatase was expressed from the parosteal layer to the endosteum, particularly in the inner osteogenic layer. ESR1 was also abundantly expressed in almost all cells from the parosteal layer to the endosteum. ARs, aromatase, and ESR1 were all abundantly expressed during the neonatal period in the hyaline cartilage of the proximal segment and in fibrocartilage of the distal segment of the baculum. Expression in all the tissues was attenuated in an age-dependent manner and became quite weak at puberty. To determine the effect of estrogen on the growth of the baculum, the aromatase inhibitor 1,4,6-androstatrien-3,17-dione (ATD) was subcutaneously injected daily into pregnant rats from Days 19 to 23 of gestation and into pups on postnatal Days 1, 3, 5, 7, and 9. On Day 10, the length of the baculum in the ATD-treated rats was significantly shorter than that in the controls, although the body weight did not change. These findings suggest that not only androgen but also locally aromatized estrogen is involved in the early growth and development of the baculum.

Ceramic Powder Made from Chicken Feces: Anti-Viral Effects Against Avian Influenza Viruses

Abstract Ceramic powder prepared by sintering of chicken feces, when mixed with avian influenza viruses or an avian adenovirus, inactivated these organisms to below detection levels. When the ceramic powder was mixed with double-distilled water, the pH of the water rose to 10 but the aqueous phase did not show any antivirus activity. After 10 washings with water or five washings with 1M Tris-HCl (pH 8.0), the ceramic powder still retained antivirus activity. Antivirus activity was not affected by the presence of organic material (33% fetal calf serum). When chicks were fed food containing 5% ceramic powder, there was no difference in body weight between normal feeding and the ceramic-mixture feeding. The mode of action of the ceramic powder remains unknown, but it possibly works by adsorbing the virus. These results show that the ceramic powder has antiviral activities and is a potentially useful tool against avian influenza on poultry farms.